Although the adult mammalian myocardium exhibits a limited ability to undergo regenerative growth, the intrinsic renewal rate is insufficient to reverse pathophysiologic cardiomyocyte loss. The ability to reconstitute lost cardiac mass in injured hearts could thus be of considerable therapeutic value. One approach to accomplish this entails inducing cell cycle activity in the surviving cardiomyocytes. Preliminary data indicate that targeted expression of cyclin D2, a key member of the regulatory complex which drives transit through the G1/S cell cycle check-point, is sufficient to induce cardiomyocyte cell cycle activity in adult hearts. Moreover, cyclin D2-induced cell cycle activity can reverse structural damage and restore function following myocardial injury. The experiments proposed in this application will further elucidate the mechanism of cyclin D-mediated cardiomyocyte cell cycle regulation. Specific Aim 1 will test the hypothesis that post-translational modification of the D-type cyclins regulates their ability to mediate regenerative growth following myocardial injury. These experiments will utilize newly generated transgenic mice expressing D-type cyclins carrying the relevant phospho-mimetic and non-phosphorylatable amino acid residue substitutions. Other studies in Aim 1 will test the hypothesis that specific phosphorylation events mediate the interaction of D-type cyclins and the p193/Cul7 E3 ubiquitin ligase, and that blocking this interaction enhances cell cycle activity in injured hearts. Experiments proposed in Specific Aim 2 will further test the hypothesis that cyclin D2-mediated cardiomyocyte cell cycle activation can be used to promote myocardial repair in the adult heart. Initial experiments will utilize a conditional transgenic mouse model to determine if cyclin D2 can induce de novo cardiomyocyte proliferation in adult hearts. Other studies are proposed to determine the degree to which pharmacologic interventions which limit adverse post-injury remodeling are able to benefit long-term, cardiomyocyte cell cycle-induced regeneration. Collectively, these studies will help establish the mechanism by which D-type cyclins regulate cardiomyocyte cell cycle entry, and the degree to which targeted expression of cyclin D2 is able to promote myocardial regeneration. The overall goal is to gain an understanding of how cell cycle regulatory pathways can be manipulation to promote the repair of injured hearts. Identification of such molecular targets may ultimately lead to the development of pharmacologic agents to promote regenerative growth in diseased hearts.